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Characterization of the cellular binding site for the urokinase-type plasminogen activator

Estreicher, A.
Schleuning, W. D.
Published in The Journal of biological chemistry. 1989, vol. 264, no. 2, p. 1180-1189
Abstract Human urokinase-type plasminogen activator (uPA) binds rapidly and with high affinity to a number of human cell types; this localizes plasmin generation to the close environment of the cell surface. uPA binding to HeLa and U937 cells is mediated by a single class of sites with an affinity of 3.4 +/- 1.3 x 10(-10) M. Binding is abolished by treatment of the cells with trypsin. Chemical cross-linking of Mr 55,000 125I-uPA to the surface of HeLa and U937 cells with disuccinimidyl suberate or with formaldehyde results in the formation of a labeled complex of Mr 100,000, suggesting a Mr of 45,000 +/- 5,000 for the receptor or a subunit thereof. When cells solubilized in Triton X-114 are subjected to heat-induced phase separation, unoccupied receptor, receptor-bound 125I-uPA, and cross-linked 125I-uPA-receptor complex all partition in the detergent phase, whereas the unbound ligand remains in the aqueous phase; similar phase partitioning is observed with endogenous uPA-receptor complexes from cultured human and murine cells. Thus, uPA bound at the cell surface is tightly associated with an amphiphilic membrane protein. Interaction of uPA with this plasma membrane receptor is species-specific, since human uPA fails to bind to murine cells, and murine uPA does not bind to human cells. Finally, incubation of HeLa cells in the presence of epidermal growth factor or phorbol 12-myristate 13-acetate results, over a period of 24 h, in a progressive change in uPA binding: an approximately 10-fold increase in the number of sites is accompanied by a 10-fold decrease in their affinity. Cross-linking and phase partitioning of 125I-uPA bound to epidermal growth factor- or phorbol 12-myristate 13-acetate-treated cells indicate that, as in control conditions, it is associated with a Mr 45,000 cell surface amphiphilic polypeptide.
Keywords Cell LineCross-Linking Reagents/metabolismEnzyme Precursors/ metabolismEpidermal Growth Factor/pharmacologyHela Cells/metabolismHumansKineticsMacromolecular SubstancesMolecular WeightPlasminogen Activators/ metabolismReceptors, Cell Surface/drug effects/isolation & purification/ metabolismReceptors, Urokinase Plasminogen ActivatorUrokinase-Type Plasminogen Activator/ metabolism
PMID: 2536017
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Other version: http://www.jbc.org/content/264/2/1180.full.pdf
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ESTREICHER, A. et al. Characterization of the cellular binding site for the urokinase-type plasminogen activator. In: Journal of Biological Chemistry, 1989, vol. 264, n° 2, p. 1180-1189. https://archive-ouverte.unige.ch/unige:8739

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Deposited on : 2010-07-12

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